Chemistry Reference
In-Depth Information
presents crystallic structure of cation of Cys obtained by X-ray diffraction [5]. Com-
plexes were introduced into erythrocyte suspension from water solution (TNIC, Cys)
or from dimethyl sulfoxide (DɆSO) solution (BTz, Pym, and Mim). The solutions
were prepared directly before experiment. The weight DMSO content in samples do
not exceeded 3%.
11.2.2 Red Cell Preparation
Mouse blood was drawn into test tube containing 3. 8% sodium citrate solution. Blood
was centrifugated for seven minute on 1,500 g. Plasma and white cells were removed
and red cells were washed three times with phosphate buffered saline (0. 85% NaCl,
5 mM NaH
2
PO
4
/Na
2
HPO
4
, pH 7. 4). Red cell mass obtained after last centrifugation
was kept at 4°C not more than 36 hr.
11.2.3 Erythrocyte Hemolysis
In experiments, 0.2% erythrocyte suspension was used. It was prepared through dilu-
tion of initial red cell mass by phosphate buffered saline on 450 times. Cell content in
the suspension amounted 4.4 × 10
7
cell/ml. Hemolytic experiments were carried out
at 37°C in plastic tubes at continuous slight stirring by magnetic stirrer. The course
of hemolysis was registered spectrophotometrically on reducing the optical density
(OD) of the erythrocyte suspension at 700 nm. At used level of the suspension dilution
(0.2% of hematocrite) the OD of suspension is lineally depended on the concentration
of undestroyed cell [6, 7]. The extent of hemolysis (Ȗ) was determined by:
A
−
A
γ
=
0
(1)
A
−
A
0
O
2
A
where A
0
and A are ODs of control and experimental samples, respectively;
is
O
2
OD of the completely hemolyzed sample.
11.2.4 Methemoglobin Determination
The 0.4 ml aliquots were removed from erythrocyte suspension and diluted 1.4 ml
distilled water. After 1 min. incubation the OD at 630 nm was measured. The methe-
moglobin content was determined by:
Δ
A
[
HbFe
3
+
]
=
630
×
d
(2)
ε
−
ε
met
oxy
where ǻA
630 =
increasing of OD at 630 nm; İ
met
= 3.8 ɦɆ
-1
sm
-1
ɢ İ
oxy
= 0.11 Mm
-1
sm
-1
molar extinction coefficients of methemoglobin and oxyhemoglobin at 630 nm, re-
spectively [8]; d = factor of dilution.
11.3 DISCUSSION AND RESULTS
The influence of SNICs on the erythrocyte hemolysis was studied in the wide con-
centration range. All of the studied complexes displayed considerable concentration
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